广谱降解有机磷农药的真菌酶解研究  

A Broad-Spectrum Organophosphorus Hydrolase from Fungus

作　　者：刘阳[1] 刘玉焕[1] 李方[1] 陈志仕[1] 廉婕[1] 罗维佳[1] 黄晓[1] 钟英长[1] 

机构地区：[1]中山大学生命科学学院生物防治国家重点实验室,广东广州510275

出　　处：《中山大学学报（自然科学版）》2004年第2期76-80,共5页Acta Scientiarum Naturalium Universitatis Sunyatseni

基　　金：广东省科技计划资助项目(C32402);广东省自然科学基金资助项目(031618);中山大学生物防治国家重点实验室开放资助项目(0203)

摘　　要：从农药污染土壤中分离出的一株真菌具有广谱降解有机磷农药的能力。菌体培养物经硫酸铵分级沉淀、SephadexG＿100凝胶过滤、DEAE＿SephroseCL6B柱层析纯化广谱有机磷农药水解酶。研究结果表明该酶为胞内酶,主要定位在细胞膜上。酶的最适反应温度为45℃,最适pH7 5,在50℃以下以及在pH6～9 5范围内活性稳定。Hg2+、Fe3+、对氯高汞苯甲酸、碘乙酸和N＿乙基马来酰亚胺对该酶有强烈的抑制作用,而Cu2+、巯基乙醇、二硫苏糖醇、二硫赤藓糖醇、谷光甘肽和去污剂对酶有不同程度的激活作用。该酶不仅可以作用于含PO键的有机磷农药;而且也能水解含PS键的有机磷农药。以甲基对硫磷和内吸磷为底物的Km值分别为63μmol L,256μmol L;Vmax分别为306μmol·min-1,189μmol·min-1;Kcat分别为1102s-1,610s-1。A fungus strain F1 isolated from the polluted soil was capable of degrading various organophosphorus pesticides effectively. A broad_spectrum organophosphorus hydrolase from mycelia of strain F1 was purified by ammonium sulfate precipitation, Sephadex G_100, DEAE_Sepharose CL_6B chromatography. The results showed that the optimal temperature and pH of the enzyme activity were 45 ℃ and pH 7.5 respectively. The enzyme was stable over the range of pH 6.0～9.5 and below 50 ℃. The activities were strongly inhibited by Hg^(2+), Fe^(3+), ρ_chloromercuribenzoate, iodoacetic acid and N_ethylmaleimide, while Cu^(2+), β_mercaptoethanol,dithiothreitol, dithioerythritol, glutathione and detergents activated the enzyme slightly. The enzyme can not only split organophosphorus pesticides possessing \{P\}\{O\} linkage such as methyl parathion, but also operate on organophosphorus pesticide containing \{P\}\{S\} bond including demeton_S. The Km, V(max), and K(cat) was 52 μmol/L, 317 μmol·min^(-1), 1 152 s^(-1) when methylparathion was as substrate, and the parameters were 236 μmol/L, 179 μmol·min^(-1), 650 s^(-1) respectively, when demeton_S as substrate.

关 键 词：广谱有机磷农药水解酶 真菌 降解 基本性质 

分 类 号：Q939.9[生物学—微生物学]